MUTATION SPECTRUM IN THE LACI GENE, INDUCED BY GAMMA-RADIATION IN AQUEOUS-SOLUTION UNDER OXIC CONDITIONS

Irradiation of DNA in a cellular environment leads to many types of DN A damage, resulting from various effects of gamma-radiation. One of th ese effects is the formation of water-derived radicals (e.g., OH radic als), which are formed in the vicinity of DNA (indirect effect). To st udy the influence of the indirect effect on gamma-radiation-induced mu tations, a newly constructed plasmid, containing the lad gene as a tar get gene, was irradiated with Co-60 gamma-radiation in aqueous solutio n, in the presence of oxygen. Under these circumstances, only OH radic als will be responsible for the induced mutations. Sequence analysis o f the gamma-radiation-induced mutations showed that 96% of all mutatio ns were base pair substitutions, 87% of which occurred in the lad gene , the others are formed in the lac operator part. All gamma-radiation- induced mutations in the lad gene occurred exclusively on G:C base pai rs, and no mutations at A:T base pairs could be detected. In the spont aneous mutation spectrum, 83% of all mutations were base pair substitu tions, 35% of which occurred in the lacI gene and 48% in the lac opera tor part. Base pair substitutions on G:C base pairs were very similar in the gamma-radiation-induced and in the spontaneous mutation spectru m, implying a high contribution of OH radicals to spontaneous mutagene sis. A:T to G:C transitions accounted for 10% of all spontaneous base pair substitutions in the lad gene and are probably the result of effe cts, other than just OH radicals. It can be concluded that OH radicals are an important source for mutations at G:C base pairs. In this pape r, the extracellular gamma-radiation-induced mutation spectrum is also compared to the previously obtained, intracellular gamma-radiation-in duced mutation spectrum of the lad gene. Comparison shows some differe nces, such as relative high amounts of mutations at A:T base pairs, G: C to T:A transversions and frameshift mutations in the intracellular g amma-radiation-induced mutation spectrum, as compared to the extracell ular gamma-radiation-induced mutation spectrum. Since the extracellula r gamma-radiation-induced mutation spectrum shows that OH radicals are mainly responsible for base pair substitutions on G:C base pairs, mut ations at A:T base pairs in the intracellular gamma-radiation-induced mutation spectrum are apparently the result of additional or other fac tors. (C) 1998 Elsevier Science B.V. All rights reserved.